Herpes simplex virus (HSV-1) is an important human pathogen responsible for self-limiting mucocutaneous lesions in immunocompetent patients and potentially lethal infections in neonates and other immunocompromised individuals. In this proposal we will test the hypothesis that HSV utilizes recombination-dependent replication to replicate its genome using both viral and cellular proteins. We propose that HSV has evolved to interact with the cellular repair and recombination machinery which the cell normally uses to respond to DMAdamage and other stress factors. The cellular machinery designed to monitor and repair damaged DMA is essential for maintaining genomic stability. Mammalian cells exposed to DNA damaging agents induce cell cycle checkpoints and DMA repair pathwaysthat serve to protect the cell from mutations and genomic rearrangements. Defects in these pathways lead to diseases such as cancer. Herpesviruses have coevolvedwith their hosts and developed fascinating ways of side stepping, subverting and in some cases benefiting from host cell responses. In this proposal we will test the hypothesis that HSV uses the homologous recombination (HR) repair pathway for its own benefit. Our preliminary work suggests that HSV uses a combination of viral and cellular proteins to carry out recombination-dependent replication needed to generate progeny genomes. We have previously demonstrated that the viral 5' to 3' exonuclease, UL12 and the major single strand DNA binding protein, ICP8, can function as a two-subunit recombinase. We propose to continue our studies of this viral recombinase and to test the hypothesis that HSV viral and cellular pathwaysfor replication of its genome. Aim 1will test the hypothesis that recombination is essential for productive viral infection; Aim 2 will test the hypothesis that ICP8 and UL12 work together during infection; and Aim 3 will test the hypothesis that cellular recombination proteins are recruited to and interact with viral preprelicative sites and active replication centers. A combination of genetic, biophysical, biochemical and cell biological approaches will be used. HSV-1 is a major human pathogen, and little is known about the mechanism of DNA replication. We have proposed that the virus utilizes some of the same machinery that cells use for preventing genetic instability and cancer. Since DNA replication is a major target for antiviral drugs, it is important to fully understand the key players in this process in order to develop better strategies for treatment. In this proposal we will test the hypothesis that HSV uses a combination of viral and cellular proteins to carry out DNA replication.